Centrifuge reagent delivery method

ABSTRACT

A device for separating components, such as fibrin I from blood, by way of centrifugation about a central axis of rotation comprises a first annular chamber defined by an outer cylindrical wall and an inner cylindrical wall, both walls being concentrically accommodated about the axis of rotation, and by a top wall and a bottom wall, where the top wall or the bottom wall is formed by a piston body displaceable within the first chamber. The device comprises furthermore a second chamber communicating with said first chamber through a first conduit and being defined by an outer cylindrical wall concentrically accommodated about the axis of rotation, said bottom wall of the first chamber, and another bottom wall. The second chamber is adapted to be placed below the first chamber during the centrifugation. The device comprises also blood feeding means for feeding blood to the first chamber and composition feeding means for feeding composition promoting the separation as well as receiving means for the connection of at least one liquid-receiving container, where the receiving means communicate with the second chamber through a second conduit. A capsule is accommodated in the second chamber and comprises a plurality of compartments for receiving respective compositions promoting the separation. The capsule comprises closing means said compartments and while influenced by the piston being adapted in sequence to open for the release of the contents of the compartments.

This is a divisional of application Ser. No. 08/566,195, filed Dec. 1,1995, now U.S. Pat. No. 5,830,352 which is a continuation-in-part ofapplication Ser. No. 08/349,166, filed Dec. 2, 1994 now abandoned.

TECHNICAL FIELD

The invention relates to a centrifuge device for separating a component,such as fibrin monomer from plasma, said method involving treating ofplasma with one or more reagents wherein said reagents are delivered toa suitable reaction chamber containing said plasma by a novel reagentdelivery system.

BACKGROUND ART

EP-PS No. 592,242 describes methods and compositions for a completelynovel fibrin sealant involving contacting a desired site with acomposition comprising fibrin monomer and converting this monomer to afibrin polymer concurrently with the contacting step. The term "fibrin"is defined as fibrin I, fibrin II, and/or des ββ fibrin.

Further a method is known from EP 0 654 669 for separating a component,such as fibrin monomer from blood. This method for separating thecomponents of a liquid containing several components of a varyingspecific gravities involves the steps of the blood being collected in afirst chamber of a device, said chamber being defined by a substantiallyaxially symmetrical outer and inner wall. The blood is subjected to acentrifugation by way of rotation of the device about the axis ofsymmetry of the chamber so as to establish a concentric interfacebetween the components of the blood. At least one of the components ofthe blood such as plasma is subsequently transferred to a second chamberin the device preferably by way of reduction of the volume of the firstchamber during a continued centrifugation of the device. Thesubstantially axially symmetrical inner wall is provided in the firstchamber so as to ensure that all the blood is subjected to a centrifugalrotation necessary for the separation. This inner wall is of a radiusadapted to the desired speed of rotation.

In the second chamber a fraction with non-crosslinked fibrin polymer isseparated from the plasma by means of a suitable enzyme and subsequentlyredissolved into fibrin monomer and transferred to a syringe through afilter by reducing the volume of the second chamber.

It turned out, however, that the separation of a component, such asfibrin monomer from blood, only by way of filtration in a device of theabove type does not provide a satisfying result. This is mainly due tothe fact that it is difficult to ensure a satisfying separation of thefraction containing fibrin monomer in the second chamber, andaccordingly a relatively high amount of the content in the blood offibrin is lost during the following transfer of a fluid fraction fromthe second chamber to the first chamber during the succeeding step ofthe method.

Also, in the earlier fibrin monomer method, the above-describedtreatment of fibrinogen within the plasma with a suitable enzymeproduced the non-crosslinked fibrin polymer in the form of a thick gelmass at the bottom of the second chamber. To provide the desired fibrinmonomer solution, a significant amount of redissolving buffer combinedwith substantial agitation was required. This resulted in severaldrawbacks. First, preferred fibrin monomer methods, e.g., for use as afibrin sealant as in EP 592,242, require concentrated fibrin monomersolutions, and the large amount of redissolving buffer or solventrequired to dissolve the gel mass provided dilute solutions which do notwork as well. Further, the substantial agitation required to dissolvethe gel mass into a fibrin monomer solution can cause damage to thedevice and to the fibrin itself.

A co-pending application entitled "Method and Device for Separating aComponent Such as Fibrin I From Blood Plasma," issued as U.S. Pat. No.5,824,230 filed concurrently herewith discloses an invention including amethod which provides for the separation of non-crosslinked fibrinpolymer from a plasma fraction in a cylindrical chamber carried outduring centrifugation whereby the non-crosslinked fibrin polymer isdeposited on the outer wall of the chamber, whereafter the remainingfluid fraction collected in the chamber is removed from the chamber, andthat the fraction with non-crosslinked fibrin polymer remaining in thechamber substantially deposited on the wall is caused to be dissolved byaddition of a solvent and by centrifugal agitation.

Since the treatment of the plasma with the enzyme is carried out duringcontinued centrifugation, the centrifugal force upon the resultingnon-crosslinked fibrin polymer provides that it is precipitated as athin gel film which substantially sticks to the circumferential walls ofthe chamber. The remaining plasma liquid deposits at the bottom of thechamber when the centrifugation is stopped and can be removed by anyconvenient means. The desired fibrin monomer solution is thereafterprovided by introducing a suitable redissolving buffer solution into thechamber and subjecting the buffer in the gel-coated chamber tocentrifugal agitation. This method provides advantages over priormethods. First, the redissolving of the non-crosslinked gel by thebuffer solution is extremely efficient due in part to the large surfacearea of the same volume of fibrin gel compared to the fibrin gel massprovided in prior methods. Accordingly, the gel can be dissolved withsmall amounts of redissolving buffer resulting in a desirablyconcentrated fibrin monomer solution. Further, the action of thecentrifugal agitation on the buffer solution within the gel-coatedchamber is a comparatively gentle method causing no damage to theequipment or to the fibrin monomer product. The resulting highconcentration fibrin monomer solution is in the range of 10-30 mg offibrin monomer per ml of solution and preferably about 25 mg/ml.

The copending invention also includes a method involving feeding ofblood preferably in the presence of an anticoagulant to a first annularchamber in a device, where the annular chamber is defined by acylindrical outer wall and a cylindrical inner wall, both wallsextending coaxially about a common axis, as well as by a top wall and abottom wall, where the top wall or the bottom wall is formed by a pistonbody displaceable within the first chamber, said method furtherinvolving a centrifugation of the device about the said common axis tosubstantially separate blood into a cell fraction and a plasma fractionfollowed by the resulting plasma fraction being transferred whileinfluenced by the piston body to a second chamber defined by an outercylindrical wall, which extends coaxially with said common axis, wherebya fraction with non-crosslinked fibrin polymer is caused to be separatedin the second chamber while a suitable enzyme is being added. Thismethod is characterized in that the fibrinogen-containing plasmafraction is subjected to the enzyme during centrifugation so that theresulting non-crosslinked fibrin polymer is deposited on the cylindricalouter wall of said second chamber, whereafter the fluid fractioncollected at the bottom of the second chamber is transferred whileinfluenced by the piston body to the first chamber, and that thefraction with non-crosslinked fibrin polymer remaining in the secondchamber substantially deposited on the cylindrical wall is caused to bedissolved by addition of a solvent and by centrifugal agitation.Thereafter the enzyme can be removed, if desired, and the so-producedfibrin monomer solution is transferred to any desired receivingcontainer.

Accordingly, an aseptic condition for collecting the solution is easilymaintained. After the fibrin monomer has been redissolved, it can betransferred to a receiving container, such as a syringe for further useas described in the prior art. Before the transfer, the enzyme can beremoved by any convenient means.

The above copending application further discloses a device forseparating components from a liquid by way of centrifugation about acentral axis of rotation comprises a first annular chamber defined by anouter cylindrical wall and an inner cylindrical wall, both walls beingconcentrically accommodated about said axis of rotation, as well as by atop wall and a bottom wall, where the bottom wall is formed by a pistonbody displaceable within said first chamber, said device furthercomprising a second chamber communicating with the first chamber througha first conduit and being defined by an outer cylindrical wallconcentrically accommodated about the axis of rotation and by saidpiston body and a bottom wall, where said second chamber is adapted tobe positioned below the first chamber during the centrifugation, andwhere said device also comprises blood feeding means for feeding bloodto the first chamber and composition feeding means for feedingcomposition promoting the separation as well as receiving means for theconnection of at least one liquid-receiving container, where thereceiving means communicate with the second chamber through a secondconduit. In a preferred embodiment, the piston rod comprises the innerwall of the first chamber.

This inventive device for carrying out the method according to thecopending invention is characterized in that the first conduit comprisesat least one channel extending between an opening at the top wall of thefirst chamber and an opening at the bottom wall of the second chamber.

As a result a device is provided which is relatively simple and whichindependent of the position of the piston ensures an easy and fasttransfer of the fractions in question from one chamber to the otherchamber, and especially of the fluid fraction from the second chamber tothe first chamber after the separation of the fibrin-I-containingfraction. The latter is especially due to the fact that the fluid isautomatically concentrated at the bottom of the second chamber when thecentrifugation is stopped, whereby it can be easily transferred to thefirst chamber by the piston being moved.

According to the copending invention it is particularly preferred thatsaid at least one channel extends through the interior of the outercylindrical wall in both the first and the second chamber with theresult that the device is particularly simple and easy to manufacture.

Further, the opening of the channel at the bottom wall of the secondchamber may be centrally accommodated in the chamber in connection witha recess formed by the bottom wall. As a result, the fluid fraction inquestion is easily and quickly guided directly to the inlet opening ofthe channel. Alternatively, each channel may be formed by a pipeextending rectilinearly through the piston body and being secured at theends in the top wall of the first chamber and the bottom wall,respectively, of the second chamber where it communicates with channelportions ending in the respective chamber.

Additionally, the first and the second chamber may in a particularlysimple manner comprise a common outer cylindrical wall shaped by anouter and an inner cylinder sealingly fitting within one another anddefining therebetween an axially extending channel, and the cylindersmay be terminated at one end by an end wall comprising an openingallowing passage of a piston rod connected to the piston body, saidpiston body forming the bottom wall of the first chamber and separatingsaid first chamber from the second chamber, and where the channelextends between the end walls of the cylinders to an opening immediatelyadjacent the piston rod.

In using such a device and method, suitable reagents for facilitatingthe separation and treatment of desired components within the bloodplasma were preloaded into the second chamber. For example, EP 592,242describes that the biotin-avidin capture system can be conveniently usedto remove the batroxobin from the desired solution. It is required thatthe biotin batroxobin be present in the second chamber to react with thefibrinogen within the plasma and convert it to a fibrin monomer (whichimmediately converts to a fibrin polymer), In order to thereaftercapture the biotinylated batroxobin using the biotin-avidin system,avidin which is bound, for example, to agarose must also be present inthe second chamber. In a closed, automated centrifuge device, theseagents need to be loaded into the device prior to blood processing.Preloading of the biotinylated batroxobin and avidin agarose into thesame chamber has provided difficulties since the high affinity of theavidin for the biotin, which is relied upon for enzyme capture, preventssufficient quantities of the enzyme from first reacting with thefibrinogen as is required.

SUMMARY OF THE INVENTION

The object of the invention is to provide a device which renders itpossible to easily place one or more reagents inside a reaction chamberand to release such reagents in a desired sequence. Preferably when usedin a device of the type described in the aforementioned copendingapplication, reagents such as an enzyme and an enzyme-capturecomposition can be released as desired. In satisfaction of the foregoingobject there is according to the invention provided a device which ischaracterized in that a capsule is accommodated in the second chamberand comprises a plurality of compartments for receiving respectivecompositions promoting the separation, and that the capsule comprisesclosing means closing said compartments and while influenced by thepiston being adapted in sequence to open for the release of the contentsof the compartments.

Such a capsule renders it possible in a simple and easy manner to feedthe substances necessary for the separation of fibrin monomer, saidcapsule preferably being provided with these substances in advance. Inaddition, the provided compartments allow a uniform predeterminedapportion of the amount in question. The batroxobin is preferably placedin one compartment in chemical relationship with biotin providing thatthe enzyme batroxobin can be easily captured after the use by means ofavidin, which is therefore placed in the second compartment in chemicalrelationship with agarose in form of relatively large particles. Thehigh affinity of the biotin for the avidin provides that complexedbiotinylated batroxobin/avidin agarose particles are subsequentlyreadily removed by filtration from the fibrin monomer solution. Theplacing of the two substances in their respective compartment renders italso possible to easily dose the substances at the desired times byinfluencing the piston. The above substances or compositions,biotin-batroxobin, respectively, and avidin-agarose can be used in anyconvenient form, e.g., lyophilized powder form.

According to the invention it is particularly preferred that the capsulecomprises a central hub coaxially mounted in the interior of the secondchamber and carrying three mutually spaced radial disks formingpartitions in the compartments and being of a substantially identicalouter circumferential contour, and that the closing means are formed bya sleeve-shaped body displaceably, but sealingly surrounding the radialdisks.

For activating the sleeve-shaped displaceable body the piston mayaccording to the invention advantageously comprise a downward skirtco-operating with the sleeve-shaped body on the capsule so as todisplace said sleeve-shaped body stepwise whereby said body in sequenceopens for release of the contents of said compartments inside thecapsule.

According to the invention the capsule may be accommodated in connectionwith an axial passage to an adjacent third chamber, the outer side ofthe sleeve-shaped body of the capsule sealingly abutting the side wallof the axial passage at least after an initial displacement of the body,whereby the lowermost partition of the capsule allows a free passage ofliquid from the second chamber to the third chamber after a finaldisplacement of the sleeve-shaped body caused by the piston out of itsengagement with the circumference of the lowermost partition. In thismanner the capsule forms furthermore part in an advantageous manner ofthe device and assist said device in its further operation during theseparation of the fibrin monomer. For the latter purpose the thirdchamber may advantageously comprise an inner annular compartment and anouter annular compartment, both compartments extending coaxially aboutthe axis of rotation, and the inner and the outer annular compartmentsmay be interconnected through a radially extending, circumferentialpassage housing an annular filter for preventing passage of liquid withsaid enzymes.

So as further to form an integrated part of the device, the hub of thecapsule may according to the invention comprise an axial, throughpassage and be secured on an upward projection centrally positioned inthe bottom of the lower, third chamber, said through passage at thebottom liquidly communicating with the outer annual compartment of thethird chamber through a channel system, and the upper end of the hub maybe adapted to be sealingly connected to an axial passage in the pistonbody so as to be connected to a liquid-receiving container securablethereto.

The methods of the present invention deal with improved processes forseparating and isolating an individual blood component or a solutioncontaining such a component. However, the present method is suitable forany procedure adaptable to a cylindrical centrifuge, wherein a firstsolution is treated with one or more catalysts or reagents duringcentrifugation. Other blood procedures which could benefit from such amethod include, but are not limited to the isolation of any bloodcomponent, such as

platelet-rich plasma,

platelet concentrate,

cryoprecipitated fibrinogen,

other proteins within plasma such as thrombin, fibronectin and the like.

Preferably the blood is from a single donor and most preferably theblood is from the same person to whom the blood component will beadministered.

While the present methods are hereinafter described in terms ofproducing a fibrin monomer solution, the scope of the invention as willbe appreciated by those skilled in the art, should not be so limited.

As used herein, the term "centrifugal agitation" refers to the motion ofthe device where the redissolving buffer solution is introduced toredissolve the intermediate product, such as non-crosslinked fibrinpolymer gel, from the outer chamber walls. Such motion or centrifugalagitation may include centrifugation to ensure that all of the exposedsurface area of the gel is subjected to the redissolving solution, andincludes preferably such a centrifugation followed by stop-and-startrotations in the same direction and/or stop-and-start rotations inopposite directions. Typical centrifugal agitations include, but are notlimited to, 5-30 second spins, preferably 5-10 second spins, at2,000-5,000 RPM in repeated forward/reverse cycles for any desiredlength of time. In the present methods, 5-10 second spins at about 3,000RPM in repeated forward/reverse cycles for 1-2 minutes is preferred. Asmentioned above, this can be preceded by a somewhat longer spin, e.g.,20 seconds or more to initially distribute the solvent.

The term "fibrin" as used herein refers to fibrin I, fibrin II or des ββfibrin.

The present device incorporating the reagent delivery system disclosedherein provides an efficient and accurate method for delivering one ormore reagents to a reaction chamber of a centrifuge. This is especiallycritical in closed, self-contained, automated centrifuges for use inblood separation techniques wherein two or more reagents are required tobe introduced into a reaction chamber in a sequential manner. In thepreferred methods and devices described herein for providing a fibrinmonomer-containing solution, e.g., for use in a novel fibrin sealant,the sequential introduction of biotinylated batroxobin followed byavidin agarose into the plasma containing chamber provides a highlysophisticated method of preparing such a solution.

In a preferred embodiment the delivery capsule also controls the fluidflow between the first and second and second and third chambers of thecentrifuge described in greater detail below and in the Figures.

BRIEF DESCRIPTION OF THE DRAWING

Preferred embodiments of the present device and methods will now bedescribed with reference to the drawing, in which

FIG. 1 is an axial, sectional view through a preferred embodiment of adevice according to the invention, and

FIG. 2 illustrates a second embodiment of the device according to theinvention.

FIG. 3 illustrates a third embodiment of the device according to theinvention.

The present device is a single, closed automatable device capable ofconverting whole blood into desired blood components preferablyautologous components useful, for example, as fibrin sealants.

The device is conveniently used within a drive unit which can secure andalign the device, rotate the device about its axis as required andactuate pistons and pushrods which will be understood to facilitate themovement of the piston, etc., from the description herein.

DESCRIPTION OF PREFERRED EMBODIMENTS OF THE PRESENT INVENTION

Preferably the present reagent delivery system is employed with a deviceas covered in the above referenced copending application and it istherefore described below with reference to such a device. However, itshould be understood that it could be employed in any reaction chamberdevice requiring delivery of one or more reagents.

The device of FIG. 1 according to the invention is built of partssubstantially presenting rotation symmetry and implying that the devicecan be placed in a centrifugation apparatus in an easy manner known perse so as to be centrifuged about a central axis 1. In this FIG. 1, apreferred embodiment of the device comprises an outer container 2 and aninner container 3 being such that they completely fit into each otherand everywhere closely abut one another apart from the portion where anaxially extending intermediary channel 4 is provided. The channel 4 isprovided by a groove shaped in the inner container 3. The two containers2 and 3 comprise their respective top portions 5 and 6, respectively,which define a central opening 7 allowing passage of a piston rod 8.About the opening 7, the two containers comprise axially extending parts9 and 10, respectively, which extend close to the hollow piston rod 8 ina direction away from the interior of the containers. The outercontainer 2 abuts the hollow piston rod along a short radially extendingflange 11 provided with a recess 12 receiving a sealing ring 13.

As illustrated in FIG. 1, the channel 4 continues between the inner andthe outer container all the way from the outer cylindrical walls of theinner and the outer container along the top portions 5, 6 and the axialparts 9 and 10 to the opening immediately below the sealing ring 13 inthe opening 7. The axial part 10 of the inner container 3 abutting theopening 7 is dimensioned such that a narrow, but free passage exists tothe interior of the containers 2 and 3 about the hollow piston rod 8.

The outer container 2 comprises a cylindrical part of a uniformdiameter, cf. FIG. 1. Downwardly when seen relative to the drawing, thispart continues into a cylindrical part 14 of a slightly larger diameterthrough a short transition part 15 forming a frusto-conical innersurface 16. The inner container 3 ends at the location where thetransition part 15 of the outer container 2 continues into thecylindrical part 14 of a larger diameter. The lower end of the innercontainer 3 comprises an outer surface 17 of a frusto-conical formmatching the form of the frusto-conical surface 16 on the inner side ofthe outer container 2. A outer and an inner annular disk 19 and 20,respectively, are provided immediately below the lower end of the innercontainer 3, which ends in a radial surface 18. These disks closely abutone another apart from the fact that they define therebetween a channel21 extending in an axial plane from a central opening 22 and forwards tothe inner side of the outer container 2, where the channel 21communicates with the channel 4 between the outer container 2 and theinner container 3 through an axially extending part 23. The channel 21and the axial channel part 23 are suitably provided by means of a groovein the side of the inner disk 20 facing the outer disk 19. The two disks19 and 20 are shaped with such an oblique course that they comprisesubstantially inner and outer frusto-conical surfaces, cf. FIG. 1, andthereby incline downwards towards the central opening 22. FIG. 1 alsoshows that the inner disk 20 comprises a radial surface 24 abutting theadjacent radial surface 18 on the inner container 3. The radial surface24 of the inner disk 20 is provided with a recess 25 for receiving asealing ring 26.

The two disks 19 and 20 are maintained in position in abutment againstthe radial surface 18 of the inner container 3 by means of a cover 27closing the outer container in the downward direction. This cover 27comprises a circumferential sleeve-shaped part 28 adapted to closelyabut the inner side of the outer container 2, to which it is secured ina suitable manner, such as by way of a snap-action by engagement betweena circumferential rib 29 on the outer side of the sleeve 28 and acorresponding circumferential groove 30 on the inner side of the outercontainer 2. A sealing connection is ensured by means of a sealing ring31 in a circumferential recess 32 at the outer periphery of the outerdisk 19. The cover 27 comprises furthermore a relatively thin wall 32adapted to form the lower bottom of the device in the position shown inFIG. 1. This wall 32 extends substantially along a course parallel tothe outer and the inner disk 19 and 20 in such a manner that the wall 32extends from the inner side of the sleeve 28 in a portion adjacent thedisks 19 and 20 and downwards towards a portion substantially on a levelwith the lower rim 33 of the outer container 2. In order to reinforcethis relatively thin wall 32, a reinforcing radial rib 34 is provided atregular intervals, only one of said ribs appearing from FIG. 1. This rib34 is shaped partly with a portion placed outside the wall 32 and partlywith a portion placed inside the wall 32, cf. FIG. 1. The latter portionis designated the reference numeral 35 and is shaped such that it abutsthe bottom side of the outer disk 19 with the result that it assists inmaintaining the disks 19 and 20 in a reliable position.

A partition means 36 is squeezed between the outer disk 19 and the cover27. This partition means 36 comprises a central pipe length 37. Thispipe length is mounted on a pin 38 projecting axially inwards and beingshaped integral with the wall 32 of the cover 27. This pipe length 37 isshaped integral with a circumferential wall disk 39 extending outwardlyfrom the pipe length 37 in such a manner that initially it inclinesslightly downwards towards the wall 32 of the cover 27 whereafter itextends along a short axial course so as to continue into a courseextending substantially parallel to the wall 32 of the cover. The walldisk 39 ends in a short radially extending periphery 40 resting on ashoulder 41 on the rib portions 35 on the cover 27. An annular filterunit 42 is squeezed between the outer periphery 40 of the wall disk 39and the bottom side of the outer disk 19. This annular filter unit 42abuts a substantially radially shaped surface 43 on the adjacent outerside of the outer disk 19. A device and methods employing such anannular filter are the subject of a copending application entitled"Centrifuge with Annular Filter" issued as U.S. Pat. No. 5,733,446.

In order to ensure a stability in the partition means 36, reinforcingradial ribs designated the reference numeral 44 are furthermoreaccommodated between the pipe length 37 and the wall disk 39.

The reagent delivery system of the present invention comprises a capsuledesignated the general reference numeral 45 is secured in the endopposite the cover 27 of the pipe length 37 of the partition means 36.Such a capsule is suitable for selectively releasing agents into thesecond chamber 75. This capsule comprises an elongated pipe length 46shaped integral with a radial ring 47 and carrying two additional radialrings 48 and 49. These radial rings 48 and 49 are secured by way ofinterference fit on their respective side of the fixed ring 47. Theloose rings 48 and 49 are accommodated at their respective distance fromthe fixed ring 47 by means of circumferential shoulders 50 and 51,respectively, on the pipe length 46. The three disks 47, 48, and 49 areall of the same outer diameter and carry along their respectiveperipheries a circumferential, displaceably mounted sleeve 52.

As illustrated in the drawing, the lower disk 49 abuts the upper end ofthe pipe length 37 of the partition means 36, whereby the position ofthe capsule 45 in the axial direction is determined. This position isfurthermore determined in such a manner that when displaced in the axialdirection the displaceable sleeve 52 of the capsule enters a sealingengagement by its lower end, cf. the drawing, with the innermost edge 53on the outer disk 19 in the central opening 22. In this position of thesleeve 52, a communication still exists between the space inside theinner disk 20 surrounding the sleeve 52 and the inlet opening to thechannel 21 between the outer disk 19 and the inner disk 20. The axiallength of the displaceable sleeve 52 is adapted such that the engagementwith the outer disk 20 occurs before the upper end, cf. the drawing, ofthe sleeve 52 disengages the fixed ring 47 during the axial downwarddisplacement of said sleeve 52. The inner diameter of the sleeve 52 isalso adapted to the outer diameter of the axially extending part of thewall disk 39 of the partition means 36 in such a manner that a continueddownward displacement of the sleeve 52 towards the cover 27 causes saidsleeve 52 to fixedly engage the partition means 36 once it hasdisengaged the outer disk 19. The length of the axial part of thepartition means 36 corresponds also to the axial length of the sleeve 52in such a manner that said sleeve 52 in the lowermost position issubstantially completely received by the partition means 36.

As illustrated in the drawing, the hollow piston rod 8 comprises acircumferential piston 55 inside the outer container 2 and the innercontainer 3, said piston 55 sealingly engaging the inner side of theinner container 3 through a sealing ring 56.

A Luer-coupling 57 is shaped inside the hollow piston rod for receivinga conventional syringe 58 with a piston-acting plug 59 for acting on thecontent of the syringe 58. The coupling 57 is shaped substantially as alength of pipe communicating with a central opening 61 in the piston 55through a frusto-conical portion 60. The length of pipe 57 is providedwith a radially inwardly projecting web 62 for directing the fluidleaving the syringe 58 away from an axial path and thereby round thelength of pipe 46 therebelow inside the capsule 45. The latter length ofpipe 46 is of such a length and such dimensions that it can sealinglyengage the length of pipe 57 inside the hollow piston rod 8 when thepiston 55 is in its lowermost position near the cover 27. In order topromote the above sealing connecting, the inner side of the length ofpipe 57 is formed with a gradually decreasing diameter at the endadjacent the piston 55.

An axially projecting skirt 63 is formed integral with the piston 55about the central opening 61 of said piston. This skirt 63 is shapedwith such a diameter and such a length that by a suitable displacementof the piston 55 it can activate the above displacement of thedisplaceable sleeve 52 of the capsule 45 into the said positions inwhich it engages the inner rim 53 of the central opening 22 through thetwo disks 19 and 20 followed by an engagement of the partition means 36.

A resilient, annular lip sealing means 64 is as indicated secured aboutthe hollow piston at the top inside the containers 2 and 3, cf. FIG. 1.This lip sealing means 64 is adapted to prevent an undesired passage offluid from the interior of the containers 2 and 3 to the channel 4, butit allows passage of fluid when a force is applied through the piston55.

As indicated at the top of FIG. 1, a connection is provided to a hose 65through an opening 66 in the outer and the inner container 2 and 3,respectively. This connection is known and therefore not shown ingreater detail, but it allows an interruption of the connection to thehose when desired. In addition, an air-escape opening with a suitablefilter is provided in a conventional manner and therefore neither shownnor described in greater detail.

A passage 69 is provided from the area between the partition means 36and the cover 27 and all the way upwards through the interior of thelength of pipe 37 of the partition means 36 and through the interior ofthe length of pipe 46 of the capsule 45. This passage 69 allows atransfer of fluid to the syringe 58 from said area when the latterlength of pipe 46 is coupled to the length of pipe 57 in the interior ofthe piston rod 8. The passage 66 is provided at the lowermost portion ofthe pin 38 in the cover 27 by said pin 38 being shaped with a plane,axial surface, said pin being of a substantially circular cross section.As a result, a space is provided between the pin and the adjacentportion of the inner side of the length of pipe 37. An area 67 isprovided immediately above the pin 38 where the partition means 36presents a slightly reduced inner diameter. In this manner it ispossible to place a small filter 68 immediately above the said area, cf.FIG. 1, whereby the fluid must pass said filter before it enters thelength of pipe 46 of the capsule 45.

The described device comprises a first annular chamber 70, which mayalso be referred to as a separation chamber, defined inwardly by thehollow piston 8 forming a cylindrical inner wall 71, and outwardly by acylindrical outer wall 27 formed by the outer container 2 and the innercontainer 3. When in the conventional use position, cf. FIG. 1, theannular chamber 70 is upwardly defined by a top wall 73 formed by thebottom 5 and 6, respectively, of the outer container 2 and the innercontainer 3. Downwardly, the annular chamber 70 is defined by a bottomwall 74 formed by the piston 55. A second chamber 75, which may also bereferred to as a reaction chamber, is defined below the piston 55, saidsecond chamber outwardly being defined by the same cylindrical outerwall 72 as the first chamber 70. Downwardly, the second chamber 75 isdefined by a second bottom wall 76 formed by the outer disk 19 and theinner disk 20. The capsule 45 is centrally accommodated in the interiorof the second chamber 75. A third chamber 77 is provided below the saidsecond bottom wall 76, and this third chamber 77 is defined by thepartition means 36 and the annular filter unit 42. In addition, thisthird chamber 77, which may also be referred to as a filtration chamber,communicates with the second chamber 75 through the passage formed bythe central opening 22 in the outer disk 19 and the inner disk 20.Finally, a fourth chamber 78 is provided below the partition means 36,said fourth chamber 78, which may also be referred to as a collectionchamber, being defined downwardly by the wall 32 of the cover 27 andfurthermore by portions of the sleeve 28 of the cover 27 and the bottomside of the outer disk 19.

As described above, the described device is primarily suited forseparation of a component, such as fibrin monomer from blood, and forthis purpose the second chamber 75, and preferably the upper chamber 80of the capsule 46, is in advance filled with a suitable enzyme, such asbatroxobin. As is understood from EP-PS No. 592,242, any thrombin-likeenzyme can be employed. Such enzymes include thrombin itself or anyother material with a similar activity, such as Ancrod, Acutin, Venyyme,Asperase, Botropase, Crotabase, Flavorxobin, Gabonase, and the preferredBatroxobin. Batroxobin can be chemically bound to biotin, which is asynthetic substance allowing the batroxobin to be captured in aconventionally known manner by means of avidin in an avidin-agarosecomposition. Accordingly, avidin-agarose is found in the lowermostchamber 81 of the capsule. Both the biotin-batroxobin composition andthe avidin-agarose composition are relatively easy to fill into therespective chambers 80 and 81 inside the capsule 45 before said capsuleis placed inside the device.

Finally, a syringe 58 is arranged, said syringe containing a pH-4 bufferprepared from an acetate diluted with acetic acid and suited forreceiving fibrin I.

Another buffer known from the prior art can also be used. Theredissolving buffer agent can be any acid buffer solution preferablythose having a pH between 1 and 5. Suitable examples include aceticacid, succinic acid, glucuronic acid, cysteic acid, crotonic acid,itaconic acid, glutonic acid, formic acid, aspartic acid, adipic acid,and salts of any of these. Succinic acid, aspartic acid, adipic acid,and salts of acetic acid, e.g. sodium acetate are preferred. Also, thesolubilization may also be carried out at a neutral pH by means of achaotropic agent. Suitable agents include urea, sodium bromide,guanidine hydrochloride, KCNS, potassium iodide and potassium-bromide.Concentrations and volumes of such acid buffer or such chaotropic agentare as described in EP-PS No. 592,242.

During or immediately after the supply of blood, the piston rod 8 ispushed so far into the interior of the device that the displaceablesleeve 52 of the capsule 45 is moved downwards into a sealing engagementin the through passage through the bottom wall 76 and to the secondchamber 77. As a result, access is simultaneously opened to thebiotin-batroxobin composition inside the uppermost chamber 80 of thecapsule.

When the device is ready for use, a blood sample is fed into the firstchamber through a needle not shown and the hose 65 in a conventionalmanner, said blood sample preferably being admixed an anticoagulant alsoin a conventional manner. During the feeding of the blood through thehose 65 and the opening 66 into the interior of the first chamber 70,air is removed from the chamber in a conventional manner. After thefeeding of blood the hose 65 is removed, and the opening 66 is sealinglyclosed. Subsequently, the device with the blood is placed in acentrifuge which inter alia assists in sealingly compressing the variousparts. The centrifuge causes the device to rotate about the axis ofrotation 1. As a result of the centrifuging, the blood is separated inthe first chamber 70 into a plasma fraction settling radially inside theremaining portion of the blood, said remaining portion containing thered and the white blood cells. As described ink EP-PS No. 592,242 theplatelets can be present in either fraction, as desired, by varying thespeed and time of centrifugation. Centrifugation speeds using thepresent device are typically in the range of 2,000-10,000 RPM and may bevaried as required at different points within the process and asdescribed herein and in EP 654 669.

When the interface between the plasma and the remaining portion of theblood has stabilized, i.e. when the separation is complete, a reductionof the volume of the first chamber 70 is initiated by the piston rod 8and consequently the piston 55 being pulled out. As a result, first apossible inner layer of air passes through the channels 4 and 21 intothe second chamber 75, and a further moving of the piston 55 impliesthat also the plasma passes to the second chamber 75. The movement ofthe piston 55 is stopped when the entire layer of plasma has been forcedinto the second chamber 75, i.e. when the interface between the plasmafraction and the remaining portion of the blood has reached the innerwall 71 of the first chamber 70.

In the second chamber 75, the plasma fraction comes into contact withthe enzyme batroxobin with the result that fibrin monomer, whichpolymerizes immediately to a non-crosslinked fibrin polymer, is releasedfrom the plasma fraction. This process is performed while the device isbeing continuously centrifuged with the result that fibrin polymer isefficiently separated from the remaining portion of the plasma fraction,said fibrin polymer being formed by the reaction of thebiotin-batroxobin composition and settling as a viscous layer along thecylindrical outer wall 72. When this separation has been completed, thecentrifuging is stopped whereby the remaining relatively fluid portionof the plasma fraction can easily be pressed back into the first chamber70 by the piston 55 first being raised for transferring air from thefirst chamber 70 to the second chamber 75 followed by said piston 55being pressed down. The fibrin polymer may remain on the outer wall ormay begin to slide off but in this case the polymer slides down muchmore slowly than the excess liquid. Thus, this transfer of fluid can beperformed relatively easily and quickly before the viscous layer withfibrin polymer reaches the opening to the channel 21. Further measurescan optionally be taken in order to prevent the viscous layer fromreaching the inlet of the channel 21 too quickly, such as by providing aring of upwardly projecting teeth 82 shown by dotted lines at the bottom76. This centrifuging/draining procedure can be carried out two or moretimes, as may be required, to get as much of the plasma fluid out of thefibrin polymer as is possible.

Once the remaining portion of the plasma fraction has been expelled fromthe second chamber 75, the displaceable sleeve 52 of the capsule 45 isfurther displaced downwards in such a manner that access is allowed tothe lowermost chamber 81. At the same time or in connection with thelatter displacement of the sleeve, the plug 49 of the syringe 58 ispressed completely downwards by means of a spindle acting from theoutside in such a manner that the pH-4 buffer is transferred to thesecond chamber 75, which can be done while initiating a centrifugalagitation. The addition of the pH-4 buffer provides that fibrin polymeris dissolved therein, and the presence of the avidin-agarose compositionin the lower chamber 81 inside the capsule 45 provides that thebiotin-batroxobin composition is bound in a conventional manner by theavidin. A continued displacement of the piston 55 causes thedisplaceable sleeve 52 on the capsule 45 to engage the partition means36 and to a disengage the bottom wall 76 with the result that a freeaccess is provided to the third chamber 77. As a result, the contents ofthe second chamber 75 can flow freely downwards into the third chamber77. Preferably, the redissolving is carried out during centrifugalagitation which involves centrifugation and a series of stop-and-startof forward/reverse agitation motions.

A continued centrifuging provides that the fibrin monomer solution canbe separated in the third chamber through the annular filter unit 42retaining the relatively large particles of agarose and the batroxobinbound thereto via the biotin-avidin capture system. When the fibrinmonomer solution has passed into the lowermost fourth chamber 78 as aresult of the above centrifuging, said centrifuging is stopped and thefibrin-I-solution is easily transferred to the syringe 58 by a renewedretraction of the plug 59, the uppermost end of the length of pipe 46 ofthe capsule 45 engaging the length of pipe 47 forming the connectionwith the syringe 58.

As fibrin polymer is separated from the plasma fraction in the secondchamber 75 during a continued centrifuging and as the fibrin monomersolution is separated in the third chamber 77 by centrifuging it ispossible to achieve a relatively high yield of fibrin monomer from theblood sample in question.

The invention has been described with reference to a preferredembodiment. Many modifications can, however, be performed withoutthereby deviating from the scope of the invention.

FIG. 2 illustrates examples of such modifications, as said FIG. 2illustrates a second embodiment of the invention which more or lesscorresponds to the embodiment of the invention shown in FIG. 1. Theembodiment of FIG. 2 comprises a first chamber 90 and a second chamber91 separated by a piston 92, which comprises a hollow piston rod 93defining the first chamber inwardly. Outwardly, the two chambers aredefined by a portion of a substantially tubular member 94 forming andouter cylindrical wall 95 for the two chambers 90 and 91. Upwardly, thefirst chamber 90 is defined by a top wall 85 which in turn is formed bya top cover secured to the tubular member 94 by means of a ring 96screwed into said tubular member 94. The top wall 85 defines a throughopening for passage of the hollow piston rod 93. Downwardly, the secondchamber 91 is defined by a bottom wall 96 formed by a circumferentialinner flange in the tubular member 94. On the side adjacent the secondchamber 91, the tubular member 94 comprises a frusto-conical surface 97inclining away from the piston 92 towards the center of the secondchamber 91. The bottom wall 96 defines a central through passage 98 to athird chamber 99. The third chamber 99 is defined by a partition means100 and an annular filter unit 101 inserted between the bottom wall 96and the partition means 100 and leading to a fourth annular chamber 102.The fourth chamber 102 is defined between a cup-shaped cover 103 securedto the tubular member 94 by threads. Said cover 103 retains throughintermediary ribs 103 the partition means 100 in position centrallyinside the tubular member 94 while squeezing the annular filter unit101.

A capsule 105 is secured on a centrally and upwardly projecting pin 104on the partition means 100. The capsule 105 comprises a tubular portion106 with disk-shaped rings 107, 108 loosely attached thereto anddefining chambers for the said enzymes indicated by the letters BB andAA, respectively, by means of a displaceably arranged sleeve. Thedisk-shaped rings are secured at the desired mutual distances on thelength of pipe 106 by means of shoulders shaped thereon by the outerperiphery of the tubular member 106 being of a decreasing diameter frombelow and upwards.

Through channels 115 and 116 are provided from the top of the firstchamber 90 to the bottom of the second chamber 91. These channels areprovided by means of their respective fixed length of pipe 117 and 118,respectively, extending parallel to the axis of rotation of the deviceand being secured at the ends in associated openings in the top wall 95and the bottom wall 96. The channel connection between these lengths ofpipe and the chambers, respectively, is provided by suitable bores andplugs secured therein. The lengths of pipe 117 and 118 extend throughtheir respective opening in the piston 92. Sealing rings are providedeverywhere so as to prevent leakage.

A coupling 120 is secured centrally inside the piston 92 for coupling toa syringe 121 inside the hollow piston rod 93 and to the upper end ofthe pipe length 106 of the capsule 105. The coupling 120 carries a skirt122 projecting into the second chamber 91 and influencing thedisplaceable sleeve 110 on the capsule 105. As illustrated, the outerdiameter of this sleeve 110 is adapted to the diameter of the throughpassage 98 downwards to the third chamber 99 in such a manner that thesleeve 110 is guided and retained by the bottom wall 96 in any positionand consequently also in a lowermost position in which the sleeve 105does not engage the lowermost disk-shaped ring 109 in the capsule andallows passage of fluid from the second chamber 91 downwards into thethird chamber 99. A channel 123 extends from the fourth chamber 102 andpasses centrally upwards through the pin 104 on the partition means 100and further upwards through the tubular member 106 of the capsule 105,whereby fluid is allowed to enter the syringe 121 therefrom.

The device of FIG. 2 is used in completely the same manner as the deviceof FIG. 1, whereby means, of course, are also provided for coupling ahose thereto for the feeding of blood.

Another embodiment is shown in FIG. 3 which has many of the same basicelements as FIGS. 1 and 2. The fluid transfer channel 4 is preferablyformed by a slot being formed within either of the inner and outercontainers 2, 3 which fit one into the other. As shown in FIG. 3, thechannel 4 extends up in between the respective top portions 5 and 6 andopens into a shoulder area 300 and does not proceed between axiallyextending parts 9 and 10 (as in FIG. 1). The resilient lip sealing means64 is directly adjacent the shoulder area 300 providing that the desiredfluid (e.g., plasma) to be transferred past the lip sealing means 64goes directly into the opening of the channel 4 between top portions 5and 6 without having to travel between the portion shaft 8 and the inneraxially extending part 10.

In another modification depicted in FIG. 3, the teeth 82 shown in FIG. 1have been replaced with a "fibrin filter" 310 which is a set of teeth ortines arranged in a circular manner (e.g., on a frame or circular ring)about the capsule 45 near the bottom of the second chamber 70. Thefilter 310 is connected in one or more locations to the bottom wall 76but is substantially open near the bottom wall 76 such that excessliquid can be drained off more efficiently. This arrangement helpsalleviate situations using the device of FIG. 1 where fibrin polymer isretained, as desired, by the teeth 82 but where excess liquid may alsobe trapped behind the fibrin polymer.

Further modifications are illustrated in FIG. 3, in particular withregards to the syringe 50 where it can be seen that a protective holder320 substantially surrounds the syringe 58. The holder 320 is preferablycylindrical or corresponds generally to the shape of the syringe 58. Aholder lid 322 is releasably attached of the syringe 58 to the top ofthe holder 320 and provides a handle for conveniently removing thesyringe 58 and holder 320 from the device after processing to providethe desired product (e.g., fibrin monomer solution) within the syringeis complete. The holder 320 can be made of a plastic or rigid polymermaterial so as to protect the syringe 58 during handling. Further, sincethe syringe is not directly touched by the operator it can betransferred to a further station for usage without contamination. Aremovable bottom lid (not shown) for the holder 320 may also beutilized, especially where the presterilized syringe 58 with holder 320and containing the required acid buffer solution is provided as onecomponent of a kit for the present device. In FIG. 3 a syringe coupler324 is also shown which is axially slidable within the lid 322 under theaction of, e.g., an upward and downward moving rod (not shown), whichmay be part of a drive unit for the device. The plug 59 is adapted toreceive the coupler 324 in both a locking and non-locking manner. Thiscan be accomplished, for example, by providing a recess 326 beyond aleveled receiving portion 328 within the interior of the plug 59 and acorresponding protuberance 330 on the shaft of the coupler 324. Thesizes and shapes of these elements are selected such that the coupler324 can be pushed downward with a minimal force to move the plug 59downward without forcing the protuberance 330 past the leveled portion320. In this way, the coupler 324 can be moved back up without changingthe position of the plug 59. If a slightly greater downward force isexerted upon the coupler 324 when engaging the plug 59, the protuberance330 will lock into the recess 326 providing that the plug 59 will nowmove in position with the coupler 324.

Also in FIG. 3, the axially projecting skirt 63 is shown to be adistinct component snugly fit within the bottom of the piston 55.

The parts described forming part of the various devices are easilymanufactured from suitable plastic materials by way of injectionmoulding, and the devices in question are therefore also relativelyinexpensive and suited for disposable use.

Accordingly, any desired materials can be used. Preferably, gammairradiatable stable polymers as are known in the medical device industryare employed. In a preferable embodiment the outer container and pistonare of polycarbonate, the syringe holder and lids and plunger are ofpolypropylene, the filter is of polyethylene, the syringe is of glass,the O-rings are of silicone and the other parts are of styreneacrylonitrile.

The invention has been described with reference to preferred embodimentsof the device. The method according to the invention may, however,easily be conducted in a laboratory under aseptic conditions by means ofa cup which can be closed by a lid. Plasma and enzyme is filled into thecup and by mixing and following centrifugation, the non-crosslinkedfibrin polymer is separated onto the bottom or the wall of the cup asdescribed above. After removing the remaining plasma fraction, thenon-crosslinked fibrin polymer is redissolved by addition of a solventand by way of centrifugal agitation as described above as well.

EXAMPLE

140 ml of whole blood and 20 ml of sodium nitrate anticoagulant (USP)was introduced into the first chamber 70 of the device described above.This combination was centrifuged for 2 minutes at about 6,000 RPM toprovide a separation of plasma and blood cells. While continuing thecentrifugation to main the separation, the piston was raised so as totransfer the innermost phase, i.e. the plasma, into the second chamber75. Approximately 60 ml of plasma was transferred. This was treated with30 units of biotinylated batroxobin which was introduced into the secondchamber 75 via the upper chamber 80 of the capsule 45 as describedpreviously. The plasma and abroxobin were mixed at a lower speed, e.g.about 2,000 to 3,000 RPM and thereafter centrifuged for 9 minutes at9,000 RPM.

The non-crosslinked fibrin polymer gel was precipitated as a thin gellayer onto the cylinder walls and the rotation was ceased. The remainingplasma fluid (serum) was then transferred back into the first chamber70. This was followed by two further 1 minute centrifugations at 9,000RPM to remove as much of the serum within the gel as possible. Followingeach such 1 minute centrifugation, the excess serum was transferred tothe first chamber 70.

Thereafter, a buffer solution comprising 3.5 ml of a 0.2M sodium acetate(pH 4.0) containing 24 mM calcium chloride was introduced into thesecond chamber 75 via the syringe 58. At this time, a centrifugalagitation consisting of 5-10 second spins at about 3,000 RPMs each inrepeated forward/reverse cycles was carried out for 2 minutes todissolve the fibrin polymer gel and provide a fibrin monomer-containingsolution. To the so-prepared solution was added avidin agarose via thelower chamber 71 of capsule 45. This was followed by a furthercentrifugal agitation consisting of 5-10 second spins at about 3,000RPMs each in repeated forward/reverse cycles for 5 minutes. Theresulting solution contained fibrin monomer plus a complex ofavidin-agarose: biotin-batroxobin.

This solution was transferred into the third chamber 77 andcentrifugally filtrated through a 20 μm annular Porex filter for 1minute at 9,000 RPM. The resulting fibrin monomer solution was collectedinto syringe 58 as described previously and contained about 25 mg/ml offibrin monomer.

The so-formed fibrin monomer solution (fibrin I in this case) wasrepolymerized into a fibrin sealant by co-administration to a site inneed of such a sealant with a 0.75M sodium carbonate/bicarbonate bufferat a ratio of fibrin I:buffer of 5:1.

I claim:
 1. A process for promoting separation of a component fromplasma comprising the steps of:feeding plasma into a closed reactionchamber comprising an outer wall, a top wall and a bottom wall; rotatingsaid reaction chamber about its longitudinal axis; opening a preloadedreagent delivery capsule within said reaction chamber by controllablysliding open an enclosing sleeve to release one or more preloadedreagents during continued rotation, said reagents being selectivelyreleased at desired times so as to promote separation of said plasmainto substantially component-free plasma and said component.
 2. Theprocess of claim 1 further comprising the step of removing thecomponent-free plasma from said reaction chamber and collecting saidcomponent.
 3. The process of claim 1 wherein said component is fibrinand at least one of said reagents is a material capable of convertingfibrinogen within said plasma into fibrin.
 4. The process of claim 3wherein the step of releasing said reagent for converting fibrinogen isdone during rotation and prior to feeding of plasma into said reactionchamber.
 5. The process of claim 3 wherein the step of releasing saidreagent for converting fibrinogen is done during rotation and afterfeeding of plasma into said reaction chamber.
 6. The process of claim 3wherein said reagent is thrombin or a substantially similar enzyme. 7.The process of claim 3 wherein the fibrin is a cross-linked polymer. 8.The process of claim 3 wherein the fibrin is a non-cross-linked polymer.9. The process of claim 3 wherein the fibrin is a monomer.
 10. Theprocess of claim 1 wherein the step of opening includes reacting saidone or more reagents and said plasma.
 11. The process of claim 10wherein said step of opening includes depositing said component on saidouter wall.
 12. The process of claim 11 further comprising the step ofadding a solvent to said reaction chamber to form a solution of thecomponent.
 13. The process of claim 12 wherein said one or more reagentsare removed from said solution of said component.
 14. The process ofclaim 13 wherein said solution of said component containing said one ormore reagents is transferred from said reaction chamber to a filtrationchamber to facilitate removal of said one or more reagents from saidsolution.
 15. The process of claim 13 wherein said solution of saidcomponent containing said one or more reagents is subjected to anotherreagent capable of binding said reagents prior to filtration so as toenhance said filtration.
 16. The process of claim 1 wherein said capsulehas a first and second reagent and wherein the step of opening includesselectively releasing a second reagent from said reagent deliverycapsule after reacting the first reagent and the plasma.
 17. The processof claim 16 further comprising the step of bonding said second reagentto the first reagent for facilitating removal of said first reagent fromsaid component.
 18. The process of claim 17 including the steps ofproviding a biotinylated biomolecule as the first reagent and whereinsaid step of selectively releasing a second reagent releases a materialhaving an affinity for biotin.
 19. The process of claim 18 wherein saidbiomolecule is thrombin or a substantially-similar enzyme.
 20. Theprocess of claim 18 wherein said second reagent is a form of avidinbound to an inert support material.
 21. The process of claim 1 whereinwhole blood is separated into a plasma fraction and a cell fraction in aseparation chamber in fluid communication with said reaction chamber andwherein said plasma fraction is thereafter fed into said reactionchamber.
 22. The process of claim 1 wherein said reaction chamber is incloseable fluid communications with a filtration chamber and whereinsaid reagent delivery capsule has first and second reagent chamberscontaining first and second reagents, respectively, said reagentchambers being enclosed by a slidable sleeve wherein(a) in a firstposition said sleeve enclosed all of said reagent chambers; (b) in asecond position said sleeve is slid downward so as to open said firstreagent chamber to release said first reagent into said reaction chamberand further where said sleeve closes said fluid communication betweensaid reaction chamber and said filtration chamber; (c) in a thirdposition said sleeve is slid further downward so as to open said secondreagent chamber to release a second reagent and continuing to close saidfluid communication between the reaction chamber and the filtrationchamber; and (d) in a fourth position said sleeve is slid furtherdownward so as to open said fluid communication between the reactionchamber and the filtration chamber.
 23. A process of promotingseparation of a component from plasma comprising the steps of:feedingplasma into a closed reaction chamber; rotating said reaction chamberabout its longitudinal axis; opening a preloaded reagent deliverycapsule within said reactin chamber to release two preloaded reagentsduring continued rotation, said reagents being selectively released atdesired times so as to promote separation of said component from saidplasma, said second reagent being selectively released from said reagentcapsule after reacting the first reagent and the plasma, said secondreagent having an affinity for the first reagent sa as to bond to andcapture said first reagent facilitating removal of said first reagentfrom said desired component.
 24. A process for promoting separation of acomponent from plasma comprising the steps of:feeding plasma into aclosed reaction chamber; rotating said reaction chamber about itslongitudinal axis; opening a preloaded reagent delivery capsule withinsaid reaction chamber to release one or more preloaded reagents duringcontinued rotation, said reagents being selectively released at desiredtimes to promote separation of said plasma into substantiallycomponent-free plasma and said component; forming a solution of thecomponent; and removing the component from the solution.